Epstein-Barr Virus (EBV) is the etiological agent of heterophile positive infectious mononuc leosis and is also associated with two malignant diseases, Burkitts lymphoma which is a common childhood cancer in tropical Africa and nasopharyngeal carcinoma, a tumor prevalent in Chinese ethnic groups. In vitro EBV is capable of transforming, "immortalizing", B-lymphocytes of human and simian origin and converting them into continuously growing lymphoblastoid cell lines which contain multiple copies of the viral genome and express the nuclear antigen EBNA. The long term goal of the proposed research is to identify the templates for those functions which mediate the transforming ability of the virus. We have previously shown that EBV DNA contains multiple copies of a 3.2kb internal repeat unit and have preliminary evidence that monkey cells transfected with the repeat sequence DNA synthesize a nuclear antigen which can be visualized using anticomplement immunofluorescence and EBV position sera. In this grant application it is proposed to utilize transfection of cloned restriction fragments of EBV DNA to: (1) Determine whether or not the nuclear antigen induced by repeat sequence DNA corresponds to the EBNA antigen. (2) Establish permanent cell lines containing the 3.2kb repeat DNA by co-selection with the HSV thymidine kinase gene. These cell lines will be used to further characterize the gene products coded by the repeat sequences and to study their regulation. (3) Define the minimal region within 3.2kb repeat unit which is required for expression of the nuclear antigen. (4) Assay for the transforming functions by screening for the ability of specific DNA fragments to induce foci of morphologically transformed cells and (5) Identify the templates for those constituents of the EBV virion against which we have, raised monoclonal antisera.